Folding of RNA molecules into specific tertiary structures is important for a large number of cellular processes relating to gene expression. For instance, protein synthesis requires transfer RNA and ribosomal RNAs to adopt functional structures, and processing of transfer and messenger RNAs depend on ribozymes or RNA-protein complexes which must also fold properly. The very high negative charge carried by RNA molecules limits their folding possibilities and causes the stabilities of folded RNAs to be very sensitive to the concentrations of Mg2+ and other ions present in solution. The long term goal of these studies is to provide a systematic and quantitative picture of ion and solvent interactions that stabilize RNA tertiary structures, and to relate the picture to the underlying electrostatic properties of RNA. A quantitative description of Mg2+ - RNA interactions that considers diffuse (hydrated) and chelated ions has been successful in accounting for the stabilities of some RNAs. To explore the limitations of this model, in future work close attention will be paid to RNAs with Mg2+ or monovalent ions in unusual environments, and the characteristics of the partially structured RNAs from which native structures fold will be studied. Other work will examine the effects of osmolytes, naturally occurring compounds accumulated by cells to high concentrations to maintain osmolarity, on RNA stability. Besides their relevance to the folding of RNAs in vivo, osmolytes may also be useful tools for characterizing the kinds of structures present in a RNA. Lastly, unusual effects of ions on the heat capacity of folded RNAs will be explored because of the potential implications for RNA hydration. Health relevance of the proposed work: Many cellular RNAs are directly involved in the synthesis of proteins, and other RNA structures regulate the levels of proteins in cells. Some of these RNAs are targets for natural antibiotics. Drugs which stabilize or alter specific critical RNA structures could have wide therapeutical applications. An understanding of the forces that stabilize RNA structures could aid in selection of drug targets and the design of new drugs.